This application claims the priority of Korean patent application Ser. No. 1999-11810 filed on Apr. 6, 1999 and Korean patent application Ser. No. 1999-47573 filed on Oct. 29, 1999.
The present invention relates to an electrochemical biosensor test strip for quantitative analysis of analytes of interest, a method for fabricating the same, and an electrochemical biosensor using the same.
In the medical field, electrochemical biosensors are extensively used to analyze biomaterials, including blood. Of them, enzyme-utilizing electrochemical biosensors are most predominant in hospital or clinical laboratories because they are easy to apply and superior in measurement sensitivity, allowing the rapid acquisition of test results. For electrochemical biosensors, electrode methods have recently been extensively applied. For example, in an electrode system established by screen printing, the quantitative measurement of an analyte of interest can be achieved by fixing a reagent comprising an enzyme onto the electrodes, introducing a sample, and applying an electric potential across the electrodes.
An electrochemical biosensor using such an electrode method may be referred to U.S. Pat. No. 5,120,420, which discloses an electrochemical biosensor test strip taking advantage of a capillary space for the introduction of analytes, teaching the use of a spacer between an insulating substrate and a cover to form the capillary space.
Another electrochemical biosensor test strip can be found in U.S. Pat. No. 5,437,999, in which a patterning technique, typically used in the PCB industry, is newly applied for the fabrication of an electrochemical biosensor, leading to an achievement of precisely defined electrode areas. This electrochemical biosensor test strip is allegedly able to precisely determine analyte concentrations on a very small sample size.
With reference to FIG. 1, there is an opposing electrode type of an electrochemical biosensor test strip described in U.S. Pat. No. 5,437,999, specified by a disassembled state in an exploded perspective view of FIG. 1A and by an assembled state in a perspective view of FIG. 1B. Typically, these sensors perform an electrochemical measurement by applying a potential difference across two or more electrodes which are in contact with a reagent and sample. As seen in the figure, the electrochemical biosensor test strip comprises two electrodes: a working electrode on which reactions occur and a reference electrode which serves as a standard potential.
There are two ways of arranging such working and reference electrodes. One is of an opposing electrode type just like that shown in FIG. 1A, in which a working electrode formed substrate is separated from a reference electrode by a spacer in a sandwich fashion. The other is of an adjacent type in which a working and a reference electrode both are fabricated on the same substrate side-by-side in a parallel fashion. U.S. Pat. No. 5,437,999 also discloses an adjacent electrode electrochemical biosensor, adopting a spacer that separates an insulating substrate, on which the electrodes are fabricated, from another insulating substrate, which serves as a cover, forming a capillary space.
In detail referring to FIG. 1, a reference electrode-formed substrate, that is, a reference electrode element 10, is spatially separated from a working electrode-formed substrate, that is a working electrode element 20 by a spacer 16. Normally, the spacer 16 is affixed to the reference electrode element 10 during fabrication, but shown separate from the reference electrode element 10 in FIG. 1A. A cutout portion 13 in the spacer 16 is situated between the reference electrode element 10 and the working element electrode 20, forming a capillary space 17. A first cutout portion 22 in the working electrode element 20 exposes a working electrode area, which is exposed to the capillary space 17. When being affixed to the reference electrode element 10, a first cutout portion 13 in the spacer 16 defines a reference electrode area 14, shown in phantom lines in FIG. 1, which is also exposed to the capillary space 17. Second cutout portions 12 and 23 expose a reference electrode area 11 and a working electrode area 21 respectively, serving as contact pads through which an electrochemical biosensor test strip 30, a meter and a power source are connected to one another.
In an assembled state as shown in FIG. 1B, the electrochemical biosensor test strip 30 has a first opening 32 at its one edge. Further, a vent port 24 in the working electrode element 20 may be incident to a vent port 15 in the reference electrode element 10 so as to provide a second opening 32. In use, a sample containing an analyte may be introduced into the capillary space 17 via either the opening 31 or 32. In either case, the sample is spontaneously drawn into the electrochemical biosensor test strip by capillary action. As a result, the electrochemical biosensor test strip automatically controls the sample volume measured without user intervention.
However, preexisting commercially available electrochemical biosensor test strips, including those described in the patent references supra, suffer from a serious problem as follows: because electrodes are planarity fabricated on substrates and reagents, including enzymes, are immobilized on the electrodes, liquid phases of the reagents are feasible to flow down during the immobilization, so that they are very difficult to immobilize in certain forms. This is highly problematic in terms of the accuracy of detection or measurement because there is a possibility that the reagent immobilized on the electrodes might be different from one to another every test strip. In addition, the electrode area exposed to the capillary space is limitedly formed in the planar substrates which the electrodes occupy. In fact, a narrower electrode area is restricted in detection accuracy.
U.S. Pat. No. 5,437,999 also describes methods for the fabrication of electrodes for electrochemical biosensor test strips, teaching a technique of patterning an electrically conducting material affixed onto an insulating substrate by use of photolithography and a technique of screen printing an electrically conducting material directly onto a standard printed circuit board substrate.
Photolithography, however, usually incurs high production cost. In addition, this technique finds difficulty in mass production because it is not highly successful in achieving fine patterns on a large area.
As for the screen printing, it requires a liquid phase of an electrically conducting material. Although suitable as electrically conducting materials for electrodes by virtue of their superiority in detection performance and chemical resistance, liquid phases of noble metals, such gold, palladium, platinum and the like, are very expensive. Instead of these expensive noble metals, carbon is accordingly employed in practice. The electrode strip obtained by the screen printing of carbon is so significant uneven in its surface that its detection performance is low.
There is also suggested a method for fabricating an electrode for an electrochemical biosensor test strip, in which a thick wire, obtained by depositing palladium onto copper, is bonded on a substrate such as plastic film by heating. This method, however, suffers from a disadvantage in that it is difficult for the electrode strip to be of a narrow, thin shape owing to its procedural characteristics. As the electric charges generated by the reaction between reagents and samples are nearer to the electrodes, they are more probable to be captured and detected by the electrodes. Hence, the bonding of a thick wire onto a plastic film brings about a decrease in the detection efficiency of the electrochemical biosensor test strip. Further, detachment easily occurs between the thick wire and the plastic film owing to a weak bonding strength therebetween and the thick electrode requires high material cost.
Therefore, it is an object of the present invention to provide an electrochemical biosensor test strip which can firmly fix appropriate reagents in a certain pattern and secure a maximal effective area of an electrode to detect charges, thereby enabling the precise quantitative determination of analytes of interest.
It is another object of the present invention to provide a method for fabricating such an electrochemical biosensor test strip, which is economically favorable as well as gives contribution to the precise detection of analytes by forming an electrode of a uniform surface.
In accordance with an embodiment of the present invention, there is provided an electrochemical biosensor test strip, comprising a first insulating substrate having a groove in a widthwise direction; a pair of electrodes parallel in a lengthwise direction on the first insulating substrate; a reagent for reacting with an analyte of interest to generate a current corresponding to the concentration of the analyte, the reagent being fixed in the groove of the first insulating substrate; and a second insulating substrate bonded onto the first insulating substrate, the second insulating substrate forming a capillary space, along with the groove.
In accordance with another embodiment of the present invention, there is provided a method for fabricating an electrochemical biosensor test strip, comprising the steps of: forming a groove in a first insulating substrate in a widthwise direction; sputtering a metal material onto the first insulating substrate with the aid of a shadow mask to form a pair of electrodes parallel in a lengthwise direction on the first insulating substrate; fixing a reagent within the groove of the first insulating substrate across a pair of the electrodes, the reagent reacting with an analyte of interest to generate a current corresponding to the concentration of the analyte; and bonding a second insulating substrate onto the first insulating substrate, the second insulating substrate forming a capillary space, along with the groove in which the reagent is fixed.
In accordance with a further embodiment of the present invention, there is provided a method for fabricating an electrochemical biosensor test strip, comprising the steps of: sputtering a metal material onto a first insulating substrate with the aid of a shadow mask to form a pair of electrodes parallel in a lengthwise direction on the first insulating substrate; fixing a reagent on the first insulating substrate across a pair of the electrodes, the reagent reacting with an analyte of interest to generate a current corresponding to the concentration of the analyte; and bonding a second insulating substrate having a groove in a widthwise direction onto the first insulating substrate, the groove being positioned across the electrodes and forming a capillary space, along with the groove, at an area corresponding to the reagent fixed.